Recurrent focal segmental glomerulosclerosis (FSGS) after transplantation is a highly prevalent condition where rituximab (a monoclonal antibody against CD20) may have a potential indication. Besides CD20, rituximab has been shown to bind sphingomyelin-phosphodiesterase-like-3b-precursor (SMPDL-3b) and to regulate acid-sphyngomyelinase (ASMase). We tested the hypothesis that rituximab prevents recurrent FSGS after transplantation via preservation of podocyte SMPDL-3b expression and activity. Our preliminary study in 41 patients at high risk for recurrent FSGS demonstrated that rituximab treatment was associated with lower incidence of post-transplant proteinuria and with decreased eGFR. The number of SMPDL-3b positive cells in post-reperfusion biopsies was reduced in patients that would later develop recurrent disease. Rituximab partially prevented the SMPDL-3b down-regulation observed in podocytes treated with the sera of affected patients. Both rituximab and SMPDL-3b overexpression prevented the disruption of the actin cytoskeleton and the apoptosis induced by patient sera, an effect that was diminished in cells where SMPDL- 3b gene was silenced. Quantitative analysis of the disruption of stress fibers in vitro was associated with the degree of post-transplant proteinuria, suggesting the possibility of developing a prediction assay for recurrent FSGS. Our University offers one of the best clinical grounds for the recruitment of a large population of patients with FSGS. We propose to utilize a pilot study in all patients with primary FSGS who undergo kidney transplantation as a feeder for the experimental studies. We will collect clinical data that will serve as secondary exploratory analyses. Post-reperfusion kidney biopsies and pre-transplant sera will be collected per protocol in patients randomized to receive rituximab or placebo. SMPDL-3b expression in kidney biopsies and in cultured podocytes exposed to patient sera will be utilized to study disease mechanisms and to perform association studies with clinical outcomes. Cell cytoskeleton remodeling, cell viability, traditional slit diaphragm proteins expression and localization and cellular lipid composition will be utilized to study if rituximab protects podocytes in a SMPDL-3b dependent manner. Our study is highly significant because it has strong clinical implications, since it may lead to a change in the approved indications for rituximab treatment of FSGS as well as other proteinuric diseases. The proposed study is innovative because it will offer new insights into the role of shyngomyelin related enzymes in podocyte function, thus allowing the identification of novel targets for antiproteinuric drug development. Finally, our in vitro assay treating normal human podocytes with patient sera may become a pre-transplant assessment test for the identifications of patients at high-risk.